Study of Hrs1 a meiosis specific component of microtubule organising centre in fission yeast S. pombe

Meiosis, the process by which gametes such as eggs and sperm are generated, is fundamental to living organisms that employ a sexual reproduction system for their propagation. The aim of sexual reproduction is to create genetic diversities, whereas the important genetic materials, which are required for essential biological activities, need to be faithfully inherited to the progenies. Meiosis is the process designed to accomplish this fiddly mission. All human cells contain two almost identical sets of genetic materials, one from the father and one from the mother. However gametes must contain only one set. This is crucial so that upon fusion of an egg and sperm, during the fertilization, the genetic content is returned to the two standard sets. Essentially, meiosis is a specialized form of cell division that halves the genetic content of parental cells. Furthermore, it allows a period of time during which one genetic set from the mother and one from the father can mix with each other leading to a unique and diverse set of genetic materials that give to the progenies their individuality. Separation of genetic materials during meiotic divisions occurs on a scaffold structure which is composed of microtubules (MTs). As their name implies, MTs are hollow rod-like tubes that are found within all cells. Specifically, they are fibrous polymers made up of the proteins alpha- and beta-tubulin, whose polymerisation and de-polymerisation enable MTs to grow and shrink. MTs contribute to the maintenance of the cell shape, the cell polarization, the cell movement and the intracellular transportation of other biological molecules and complexes. These important and diverse functions of MTs rely on their dynamic properties which, in turn, are regulated by MT-associated proteins. MTs emerge from a specialized structure within the cell called the MT organising centre (MTOC). MTOC contains another type of tubulin, gamma-tubulin, which, in association with several other proteins, forms a structure called gamma-tubulin complex (gamma-TuC) that acts as a nucleation seed for alpha/beta tubulin dimers to polymerise. In addition, many of the MT regulatory proteins are found to be concentrated at the MTOC. In this project, I aim to investigate MT regulation during meiotic cell division using yeast as a model organism. Many advances in understanding biology come from leads established in model systems. Fundamental cellular activities, such as ones performed by MTs, are conserved from yeasts to humans. A prime example of the impact of model systems upon the field is the identification of gamma-tubulin in a filamentous fungus A. nidulans. Search for homologous proteins of A. nidulans gamma-tubulin identified human gamma-tubulin. Another example is that extensive analyses employing budding yeast S. cerevisiae brought about identification and insights into conserved components of gamma-TuC. Following these leads I also aim to exploit in my work a highly tractable system the fission yeast S. pombe. In fission yeast, extensive conformational change of MT architecture takes place during meiosis. I have previously identified a protein called Hrs1, which appears on the MTOC during the early stage of meiosis, and induces a special MT conformation through this stage. I assume that there exists a regulatory system which makes Hrs1 to appear temporarily on the MTOC and assists the transition from one meiotic stage to the other. In this proposal, I will examine this assumption to obtain better insight into the MTOC regulation. Model systems are living test tubes which enable us to precisely examine working hypotheses in order to extract concepts applicable to other living organisms. Placing such information in the context of what we know about the MTOC of less tractable organisms provides valuable insights into the general MTOC function.

Dynamic changes in the conformation and properties of microtubules (MTs) is vital for many essential cell biology processes. These changes occur with the help of regulatory proteins which are often concentrated at the microtubule organising centre (MTOC) in a proliferation and/or differentiation stage specific manner. I aim to exploit a highly tractable genetic system, the fission yeast S. pombe, to study the regulation and function of the primary MTOC, the spindle pole body (SPB; the fungal equivalent of the centrosome) during meiosis. In fission yeast, extensive remodelling of MT architecture takes place during meiotic differentiation. I have previously identified a protein, Hrs1, which appears on the SPB upon commitment to meiotic differentiation, and shown that Hrs1 is responsible for the formation of an extensive astral array of MTs that is necessary for the dramatic nuclear movements associated with meiotic recombination. Hrs1 then disappears from the SPB in meiosis I when spindle formation takes place. A mutant protein, Hrs1.TA, which persists on the SPB even after the onset of meiosis I, delays spindle formation. From these observations, I postulate that downregulation of Hrs1 is necessary to assist the transition from meiotic prophase to meiosis I. I will test this hypothesis by first performing a detailed analysis of SPB structure and Hrs1 localization during meiosis. I will then analyse the molecular events involved in Hrs1 downregulation and examine the mechanism by which Hrs1.TA delays spindle formation. Finally, I will isolate interacting partners of Hrs1 that contribute to its role in MT reorganization. These experiments will shed new light on how MT dynamics are controlled during meiotic recombination and chromosome segregation. Moreover, due to the conservation of eukaryotic cell cycle processes, we expect these results to provide important insights into the regulation of meiosis, and possible mitosis, in humans.